Studies will be undertaken to investigate the mode of regulation of bovine brain and ascites tumor hexokinase. The experiments will involve initial rate studies and static binding experiments. Attempts will be made to evaluate the mode of inhibition of various sugar-phosphate analogs of glucose-6-P. It will be of interest to study the potential inhibitory action of these compounds and their reversal by orthophosphate. These investigations may provide insight into the effector structural requirements for regulation of hexokinase. Almost all of what is known regarding hexokinase function and regulation has arisen from initial rate studies. In these experiments the level of enzyme is far below the concentration found in vivo. Kinetic experiments will be carried out using hexokinase at its intracellular concentrations. These data will then be compared with results currently available on the kinetic mechanism and mode of regulation of hexokinase.